Sex differences in expression and subcellular localization of heart rhythm determinant proteins.

To evaluate sex differences in protein expression in the heart, we performed Western blot studies on a subset of Heart Rhythm Determinant (HRD) proteins. We examined key components of a variety of types of mechanical and electrical junctions including, connexin43, plakophilin-2, N-cadherin and plakoglobin, ankyrin-2 and actin. We describe novel findings in sex differences in cardiac protein expression and membrane localization. For most proteins examined, sex differences were significantly more pronounced in the membrane compartment than in overall expression. These studies extend our previous findings in microarray studies to demonstrate that sex differences in gene expression are likely to confer distinct functional properties on male and female myocardium.

Postobstructive regeneration of kidney is derailed when surge in renal stem cells during course of unilateral ureteral obstruction is halted.

Unilateral ureteral obstruction (UUO), a model of tubulointerstitial scarring (TIS), has a propensity toward regeneration of renal parenchyma after release of obstruction (RUUO). No information exists on the contribution of stem cells to this process. We performed UUO in FVB/N mice, reversed it after 10 days, and examined kidneys 3 wk after RUUO. UUO resulted in attenuation of renal parenchyma. FACS analysis of endothelial progenitor (EPC), mesenchymal stem (MSC) and hematopoietic stem (HSC) cells obtained from UUO kidneys by collagenase-dispersed single-cell suspension showed significant increase in EPC, MSC, and HSC compared with control. After RUUO cortical parenchyma was nearly restored, and TIS score improved by 3 wk. This reversal process was associated with return of stem cells toward baseline level. When animals were chronically treated with nitric oxide synthase (NOS) inhibitor at a dose that did not induce hypertension but resulted in endothelial dysfunction, TIS scores were not different from control UUO, but EPC number in the kidney decreased significantly; however, parenchymal regeneration in these mice was similar to control. Blockade of CXCR4-mediated engraftment resulted in dramatic worsening of UUO and RUUO. Similar results were obtained in caveolin-1-deficient but not -overexpressing mice, reflecting the fact that activation of CXCR4 occurs in caveolae. The present data show increase in EPC, HSC, and MSC population during UUO and a tendency for these cells to decrease to control level during RUUO. These processes are minimally affected by chronic NOS inhibition. Blockade of CXCR4-stromal cell-derived factor-1 (SDF-1) interaction by AMD3100 or caveolin-1 deficiency significantly reduced the UUO-associated surge in stem cells and prevented parenchymal regeneration after RUUO. We conclude that the surge in stem cell accumulation during UUO is a prerequisite for regeneration of renal parenchyma.

Beneficial effects of long-term use of the antioxidant probucol in heart failure in the rat.

BACKGROUND: Congestive heart failure (CHF) is a disease that is characterized by progressive left ventricular (LV) dysfunction and dilatation. Oxidative stress is thought to contribute to the progression of CHF, and antioxidants have been shown to have beneficial effects when started early after myocardial infarction (MI). In this study, we tested whether the powerful antioxidant probucol would attenuate progression of CHF once it was established after MI in the rat. METHODS AND RESULTS: Ligation of a coronary artery was used to create an MI in rats (n=266). Survivors were then randomized 20 days after MI to either probucol 61 mg. kg(-1). d(-1) or vehicle and followed up for a total of 100 days after MI. Studies of cardiac hemodynamics, LV remodeling, cardiac apoptosis and morphology, systemic neurohumoral activation, oxidative stress, and renal function were then evaluated. Probucol improved LV function (LV maximum rate of pressure rise from 3103 to 4250 mm Hg/s, P<0.05, and LV end-diastolic pressure decrease from 28 to 24 mm Hg, P<0.05), reduced pulmonary weights, prevented right ventricular systolic hypertension, and preserved renal function compared with vehicle. Probucol also prevented LV dilatation, prevented wall thinning (1.70 versus 1.42 mm, P<0.05), reduced cardiac fibrosis and cardiac apoptosis, attenuated increased myocardial cell cross-sectional area, and increased scar thickness. CONCLUSIONS: In chronic CHF, probucol exerts multiple beneficial morphological effects that result in better LV remodeling and function, reduced neurohumoral activation, and preserved renal function.

Importance of local production of endothelin-1 and of the ET(B)Receptor in the regulation of pulmonary vascular tone.

Interaction between locally released endothelin-1 (ET-1) and the endothelial ET(B)receptor could modulate pulmonary vascular tone. We evaluated pulmonary ET-1 clearance and ET-1-ET(B)receptor interaction in the modulation of pulmonary vascular tone. Controls and rats with Monocrotaline (MCT)-induced pulmonary hypertension (PH) were studied. Lungs were isolated and perfused under constant pressure. The effect of the selective ET(B)antagonist BQ-788 (10(-12)-10(-8)mole) on perfusion flow rate and(125)I-ET-1 extraction was determined. Baseline(125)I-ET-1 extraction was reduced from 62+/-5% in controls to 49+/-10% in PH (P=0.012). BQ-788 inhibited extraction with a higher half-inhibitory dose in the MCT group (-Log ID(50)= 8.9+/-0.4 vs. 9.5+/-0.1, P=0.03). BQ-788 induced a mild reduction in perfusion flow rate of 0.7+/-0.3 ml/min in controls. In the MCT group, this occurred at a lower dose and was more pronounced with a maximal reduction of 3.3+/-0.7 ml/min (P<0.01 vs. control). ET-1 was undetectable in the effluent at baseline but was present in similar concentrations in both groups after ET(B)blockade. Addition of 2 pg/ml ET-1 to lung perfusate did not modify pulmonary ET-1 clearance or the effect of BQ788 on perfusion flow rate in control lungs. In normal rat lungs, the ET(B)receptor plays a minor regulatory role on vascular tone. In MCT hypertension however, despite a reduction in ET(B)mediated extraction, luminal production of ET-1 attenuates the increase in pulmonary vascular tone.

Effectiveness of a nonselective ET(A/B) and a selective ET(A) antagonist in rats with monocrotaline-induced pulmonary hypertension.

BACKGROUND: Both nonselective ET(A/B) receptor and selective ET(A) receptor antagonists can reduce pulmonary hypertension (PH) and right ventricular hypertrophy (RVH) in various animal models. Depending on their net effects after blockade of endothelial and smooth muscle ET(B) receptors, nonselective ET(A/B) antagonists could be more or less effective than selective ET(A) antagonists. METHODS AND RESULTS: Two weeks after injection of saline or 60 mg/kg monocrotaline (MCT), rats received 50 mg x kg(-1) x d(-1) of a selective (LU135252) or nonselective (BSF420627) antagonist for 3 weeks. This resulted in 4 groups: control (n=15), MCT (n=60), MCT+ET(A) (n=39), and MCT+ET(A/B) (n=40). Five-week survival was 35% in the MCT group; this was increased to 56% in the MCT+ET(A) group (P:=0.10) and to 67% in the MCT+ET(A/B) group (P:=0.0015). Drug administration was stopped 48 hours before hemodynamic measurements to evaluate the chronic effects of therapy: PH in the MCT group (RV systolic pressure 87+/-1 mm Hg) was improved similarly in both MCT+ET(A) and MCT+ET(A/B) groups (72+/-3 and 70+/-3 mm Hg, respectively, P:<0.05). Severe RVH in the MCT group (RV/left ventricle+septum weight ratio 73+/-1%) was not affected by the selective antagonist (70+/-2%) but was reduced to 54+/-2% in the MCT+ET(A/B) group (P:<0.01). Pulmonary resistive properties, assessed from isolated lung pressure-flow relationships, were improved similarly in survivors from both treated groups. CONCLUSIONS: Both the nonselective ET(A/B) antagonist BSF420627 and the selective ET(A) antagonist LU135252 are effective in this model of PH. Similar direct comparative studies in other models of PH and with various dosage regimens are warranted to define the optimal pharmacological approach of PH when ET receptor antagonists are used.

Effect of ET(A) receptor antagonist on pulmonary hypertension and vascular reactivity in rats with congestive heart failure.

BACKGROUND: We evaluated the effects of chronic therapy with the selective ET(A)receptor antagonist LU 135252 (LU) on pulmonary vascular reactivity in congestive heart failure. METHODS AND RESULTS: After myocardial infarction (MI) or sham operation, rats were gavaged with LU or saline for 4 weeks. Studies were performed in isolated lungs and to differentiate acute from chronic effects, some experiments were performed with LU in the perfusate. The MI+saline group developed moderate pulmonary hypertension (PH) and right ventricular hypertrophy (RVH). This was improved by LU therapy despite no change in left ventricular function and a larger scar area. Vasodilation to sodium nitroprusside (SNP) was reduced after MI and modestly improved by LU therapy. Vasodilation to acetylcholine (Ach) was similar among the four groups, but accentuated after acute LU administration in the sham+saline and MI+saline groups. A23187 produced higher vasoconstriction (18+/-4%) in the MI+LU compared to the MI+saline (10+/-3%, P<0.05) and the two control groups (3+/-1% and 4+/-3%, with and without LU, P<0.01): this was reversed to vasodilation following the acute addition of LU. CONCLUSION: ET(A)receptor blockade after MI reduces PH and RVH. LU therapy mildly improves dilation to SNP and favorably modulates pulmonary endothelium-dependent responses. These results support future studies to better define the mechanisms of improvement in pulmonary vascular reactivity after ET receptor antagonist therapy.